This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Our study focuses on a non-cancerous cell line (RWPE-1) and three cancerous lines (DuCaP, VCaP, and LnCaP). Two of these prostate cancer lines (DuCaP and VCaP) represent novel cell lines derived from aggressive tumors. We have defined the total Zn2+ levels in these cells using Inductively Coupled Mass Spectrometry (ICP-MS) and demonstrate that the cancer cell lines display a marked reduction in total Zn2+ as well as an inability to concentrate Zn2+ upon treatment with exogenous metal. The main focus of our work has been to develop and use fluorescent sensors and live-cell fluorescence microscopy to image labile Zn2+ in different regions of the cell. We have discovered that there are significant differences between the cancerous and non-cancerous cells;however, we only have probes for some subcellular locations. This study would be greatly strengthened by developing a comprehensive picture of Zn localization in cells. To understand why prostate cancer cells exhibit less Zn than healthy cells, we would like to pursue measurements that enable us to define whether there is an overall homogenous reduction in Zn, or whether Zn is depleted from specific sub-cellular locations. Fortunately, micro-XRF methods have the potential for significant impact as a complement to live-cell imaging. Our goal is to map zinc levels and distribution in cancerous vs. non-cancerous prostate cells in order to define the cellular and sub-cellular changes that occur in disease progression. This is a critical first step in defining the role zinc plays in normal prostate and elucidating how zinc depletion correlates with the onset of cancer. Synchrotron-based X-ray fluorescence microprobe measurements are ideally suited to quantitatively map zinc topography &heterogeneity on an appropriate length scale in fixed cells. Additionally, this would compliment our live cell studies.